U.S. Pat. No. 5,985,848, U.S. Pat. No. 6,066,722 and U.S. Pat. No. 6,228,741 are directed to nucleoside analogues that are inhibitors of purine nucleoside phosphorylase (PNP). The analogues are useful in treating parasitic infections, as well as T-cell malignancies and autoimmune diseases.
PCT/NZ00/00048 provides a process for preparing these PNP inhibitor compounds. This application recognises the compounds as PNP inhibitors and addresses a need for simpler methods of preparing them.
PNP catalyses the phosphorolytic cleavage of ribo- and deoxyribonucleosides, for example those of guanine and hypoxanthine, to give the corresponding sugar-1-phosphate and guanine, hypoxanthine or other purine bases.
The applicants have now determined that certain of these PNP inhibitor compounds are actually powerful and biologically available inhibitors of 5′-methylthioadenosine phosphorylase (MTAP) and 5′-methylthioadenosine nucleosidase (MTAN).
MTAP and MTAN function at or near the crossroads of polyamine biosynthesis, and of purine salvage in mammals and microbes, and of quorum sensing pathways in microbes. They respectively catalyse the reversible phosphorolysis of 5′-methylthioadenosine (MTA) to adenine and 5-methylthio-α-D-ribose-1-phosphate (MTR-1P), and the hydrolysis of MTA to adenine and 5-methylthio-α-D-ribose. The adenine formed is subsequently recycled, converted into nucleotides and is essentially the only source of free adenine in the human cell. The MTR-1P is subsequently converted into methionine by successive enzymatic actions.
Scheme 1 shows the role of MTAP and MTA in polyamine biosynthesis. Scheme 2 shows the reaction catalysed by MTAP (phosphorolysis of MTA to adenine and 5-methylthio-α-D-ribose-1-phosphate) including the proposed transition state structure.


MTA is a by-product of the reaction involving the transfer of an aminopropyl group from decarboxylated S-adenosyl methionine to putrescine during the formation of spermidine. The reaction is catalyzed by spermidine synthase. The spermidine synthase is very sensitive to product inhibition by MTA, therefore inhibition of MTAP or MTAN will severely limit the polyamine biosynthesis and the salvage pathway for adenine in the cells.
Inhibition of MTAN may also decrease production of the quorum sensing pathways in bacteria, and thereby decrease the virulence of microbial infections.
In the Al-1 quorum sensing pathway, S-adenosylmethionine (SAM) and specific acyl-acyl carrier proteins are the substrates for homoserine lactone (HSL) biosynthesis. The biosynthesis of HSL results in concomitant release of MTA. Thus, a buildup of MTA due to inhibition of MTAN should result in inhibition of the Al-1 pathway.
In the Al-2 quorum sensing pathway, SAM is converted to S-adenosylhomocysteine (SAH), then to S-ribosylhomocysteine, and on via 4,5-dihydroxy-2,3-pentanedione to the Al-2 quorum sensing molecule. The SAH is a substrate for MTAN, so inhibition of MTAN should directly inhibit the Al-2 pathway.
MTAP deficiency due to a genetic deletion has been reported with many malignancies. The loss of MTAP enzyme function in these cells is known to be due to homozygous deletions on chromosome 9 of the closely linked MTAP and p16/MTS1 tumour suppressor gene. As absence of p16/MTS1 is probably responsible for the tumour, the lack of MTAP activity is a consequence of the genetic deletion and is not causative for the cancer. However, the absence of MTAP alters the purine metabolism in these cells so that they are mainly dependent on the de novo pathway for their supply of purines. That makes these cells unusually sensitive to inhibitors like methotrexate and azaserine, that block the de novo pathway. Therefore, a combination therapy of methotrexate or azaserine with an MTAP inhibitor will have unusually effective anti-tumour properties.
MTAP inhibitors are may also be effective as radiation sensitizing agents. The inhibition of MTAP could result in a reduced ability to repair damage caused by ionising radiation.
MTAP inhibitors would also be effective against parasitic infection such as malaria that infects red blood cells (RBCs). It has been shown that Plasmodium falciparum has an active MTAP pathway (Sufrin, J. R., Meshnick, S. R., Spiess, A. J., Garofolo-Hannan, J., Pan, X- Q. and Bacchi, C. Y. (1995) Antimicrobial Agents and Chemotherapy, 2511–2515). This is a target for MTAP inhibitors. Such inhibitors may also kill the parasites without having any negative effect on the host RBCs, as RBCs are terminally differentiated cells and they do not synthesize purines, produce polyamines or multiply.
The polyamine pathway is important in cancer development, and blocking the polyamine pathway with inhibitors of MTAP is expected to provide reduced growth of cancers. Genetically modified mice (TRAMP mice, Gupta, S., Ahmad, N., Marengo, S. R., MacLennan, G. T., Greenberg, N. M., Mukhtar, H. (2000) Cancer Res. 60, 5125–5133) with a propensity for prostate tumor development have been described. Treatment of these mice with known inhibitors of the polyamine pathway such as α-difluoromethylornithine (DFMO) delay the onset of cancers and prevent metastasis to other tissues. However, the use of DFMO in humans is limited by its ototoxicity (causes deafness). MTAP inhibitors target a different step in the polyamine pathway, and are also expected to reduce cancer development. Since MTAP inhibitors influence a different step in the pathway, one that is only used in the polyamine pathway in humans, they may act without the side-effects that have limited the application of other polyamine pathway inhibitors.
It is an object of the present invention to provide compounds that are inhibitors of MTAP and/or MTAN, or at least to provide the public with a useful choice.